The synthesis of new core molecules is often undertaken to provide different orientations of the attached building blocks, thereby increasing structural diversity. The properties of the core molecule have (in several assays) been critical to activity, as libraries made with the same building blocks and linkages have had very different activity levels (Carell et al. Chem. Biol. 1995, 2, 171–183). We have previously demonstrated the use of 1, 3, 5, 7-cubanetetracarboxylic acid chloride (Carell et al. Angew. Chem., Int. Ed. Engl. 1994, 33, 2059–2061; Eaton et al. Angew. Chem., Int. Ed. Engl. 1992, 31, 1421–1436; Bashir-Hashemi, A. Angew. Chem., Int. Ed. Engl. 1993, 32, 612–613) and both 9,9-dimethyl-2, 4, 5, 7-xanthenetetracarboxylic acid chloride and -tetraisocyanate as cores for combinatorial chemistry (Carell et al. Chem. Biol. 1995, 2, 171–183; Carell et al. Angew. Chem., Int. Ed. Engl. 1994, 33, 2061–2064; Carell et al. Angew. Chem., Int. Ed. Engl. 1994, 33, 2059–2061; Shipps et al. Bioorg. Med. Chem. 1996, 4, 655–657; Pryor et al. Tetrahedron 1998, 54(16), 4107–4124).
The fact that the certain receptors and proteins appear to bind their ligands utilizing small clusters of residues for the majority of the binding interaction has led to the expectation that small molecules may be capable of triggering a receptor response. It has been anticipated that the generation of detailed knowledge concerning the dimerization modes and ligand binding domains of single transmembrane domain receptors will provide a basis for the design of functional agonists as well as ligand antagonists. However, the noncontiguous and multiple binding domains involved in both the protein-protein and ligand-protein interactions make it difficult to assess the dimerization mode or ligand binding domains in the absence of three-dimensional structural information. This is especially true considering the size of the typical endogenous ligands including proteins such as EPO (166 residues) which themselves contain noncontiguous binding domains which intereact with both subunits of the dimerized receptor.
Recently, the successful identification of cyclic polypeptides with the capacity to mimic the action of EPO was reported, together with details of the intricate receptor-ligand and receptor-receptor interactions in the bound complex (Wrighton et al. Science 1996, 273, 458; Livnah et al. Science 1996, 273, 464). Although these results represent a major achievement, the size (2 to 20 residues) and nature of ligands identified would not seem to be immediately applicable as drug candidates.
In a recently published PCT application (Rebek et al. WO 95/19359), a process for making xanthene or cubane based compounds and protease inhibitors is described. More particularly, methods for forming combinatorial libraries and the libraries produced are provided. According to a preferred aspect of the invention, a plurality of core molecules, the core molecule being a xanthene or cubane derivative, are reacted with a plurality of different “tool” molecules to form a library of molecules having non-naturally occurring molecular diversity. The libraries are useful for identifying lead compounds which modulate the functional activity of a biological molecule. Protease inhibitors that have been isolated from the libraries also are disclosed.
Combinatorial chemistry, introduced for polypeptide and oligonucleotide libraries, has undergone a rapid development and acceptance. It is widely recognized that this approach, when applied to generating non-peptide small molecule diversity, has provided a new paradigm for drug discovery. Perhaps as a consequence of the extension of the concept from peptide and oligonucleotide synthesis, the majority of applications have relied on solid-phase synthesis and methodological advances continue to extend common synthetic transformations to polymer-supported versions (Thompson et al. J. A. Chem. Rev. 1996, 96, 555; Früchtel et al. Angew. Chem., Int. Ed. Engl. 1996, 35, 17; Hermkens et al. Tetrahedron 1996, 52, 4527).
A less frequently used complement to adapting solution-phase chemistry to polymer-supported combinatorial synthesis is the development of protocols for solution-phase combinatorial synthesis (Han et al. Proc. Natl. Acad. Sci. U.S.A. 1995, 92, 6419). Preceding the disclosure of efforts on the the development of a multi-step solution-phase parallel synthesis of chemical libraries (Cheng et al. J. Am. Chem. Soc. 1996, 118, 2567; Boger et al. J. Am. Chem. Soc. 1996, 118, 2109; Cheng et al. Bioorg. Med. Chem. 1996, 4, 727, Tetrahedron Paper and Patent), the single-step solution-phase synthesis of combinatorial libraries was detailed by at least three groups as follows. Smith and coworkers (Smith et al. Bioorg. Med. Chem. Lett. 1994, 4, 2821), prepared a library of potentially 1600 amides by reacting 40 acid chlorides with 40 nucleophiles. The library was screened as 80 sample mixtures in a matrix format, allowing immediate deconvolution.
A similar sub-library format was used by Pirrung and Chen (Pirrung et al. J. Am. Chem. Soc. 1995, 117, 1240; Pirrung et al. Chem. Biol. 1995, 2, 621) who prepared a series of carbamate mixtures which were screened for acetylcholinesterase inhibitory activity. Prior to these efforts, we have disclosed the single-step construction of large libraries presenting amino acid derivatives attached to rigid core templates with a reliance on amide or urea bond formation (Carell et al. Angew. Chem., Int. Ed. Engl. 1994, 33, 2059; Carell et al. Bioorg. Med. Chem. 1996, 4, 655; Dunayevskiy et al. Anal. Chem. 1995, 67, 2906; Carell et al. Chem. Biol. 1995, 2, 171). Because of the complexity of the combinatorial libraries resulting from this approach (approaching 100,000 members), an iterative selection strategy based on structural grouping of the building blocks was devised.
In addition to recent advances in this work, substantial progress towards using solution-phase multicomponent reactions for generating combinatorial mixtures has been disclosed. For example, both Ugi and Armstrong have reported four-component condensations including the incorporation of a modifiable isocyanide in combination with resin capture strategy, to provide useful solution-phase library preparations (Ugi et al. Endeavour 1994, 18, 115; Keating et al. J. Am. Chem. Soc. 1996, 118, 2574; Armstrong et al Acc. Chem. Res. 1996, 29, 123).
There remains a need in the art, however, for small molecule libraries of chemical compounds and economical methods for producing such libraries for use in protein and receptor targets as described above. Furthermore, what is needed is an economical method for the identification or deconvolution of these active chemical libraries to rapidly determine active components.